7.2.2. Core B. Instrumentation and systems integration Core Leader: Mike Mandella Investigators: Thomas Huser, Gordon Kino, Pierre Khuri-Yakub, Craig Levin, Olav Solgaard, David Paik, Dave Piston, Kenneth Salisbury Aim 1. Validation of new microscopy and establishment of standards for miniaturized intravital microscopes for preclinical and clinical use A key aspect of this program is the availability of tools for validating the new optical imaging methods for specific cancer applications. We have designed part of Core B specifically for quantitative comparisons of performance with other imaging modalities or histopathology on biopsy specimens. Such validation is necessary prior to any clinical trial designed to demonstrate the clinical efficacy of the optical methods being developed in this program. Core B will perform pre-clinical evaluation and analyses of human samples. Since in vivo optical measurements usually preserve tissue structure, verification of optical signatures should then be possible by correlation with measurements in vitro. Part of the validation will be through sharing our instruments and reagents with other programs in the Network to get external validation of our tools. We will compare and share the results from our studies on optical imaging methods, contrast agents, and software with other Network investigators to develop consensus on robust methods for validation in the target cancer applications. Aim 2. integration of microscopy into wide-field fluorescence systems and co-registration of data with ultrasound The dual-axes microscope provides high resolution images in living tissue up to 0.5 mm deep and within a field-of-view of about 0.5 mm diameter. Because of this limited field-of-view, it will be important to provide context by spatially registering these micro-scale images to the larger anatomical data acquired with both, ultrasound and wide-field fluorescence, and to develop an integrated software platform for real-time micro and macroscopic image processing. Wide-field fluorescence imaging can be obtained by commercially available endoscopes such as the Olympus FQ260Z GIF gastrointestinal videoscope. By inserting the dual-axes microscope into the provided instrument channel of such a fluorescence endoscope, we can thereby obtain meaningful microscopic fluorescence images by letting the microscope placement be guided to the best micro-imaging sites by assistance from the wide-field macro-imaging system. In addition to using the instrument channel for microscopy, we also plan to modify the wide-field fluorescence endoscope by installing a conformal ultrasound transducer imaging array around the -12 mm outer diameter of the endoscope's tubular housing. The wide-field fluorescence images will allow mapping of the surface of the esophagus and the 3-D ultrasound images will provide information about the underlying tissue up to several millimeters deep below the same surface. We will develop algorithms for co-registration of these two different types of images, which will provide precise locations of the lesion sites for guiding the placement of the dual-axes microscope as it is extended towards the tissue through the instrument channel of the endoscope. These algorithms will be developed in stages, beginning with simple registration algorithms for displaying real-time images of the three different imaging modalities (microscopy, ultrasound, and wide-field fluorescence). More advanced algorithms will then be implemented as they are developed, which will have increasing functionality, such as the addition of real-time mosaicing with the dual-axes microscope and advanced methods for displaying the data. We will work towards achieving registration of the three different types of images to an anatomical map in real-time and displayed in 3-D at their respective imaging depths. In this case, the widefield fluorescence image data would be displayed at the surface, with microscopy data extending deeper, and finally with the ultrasound data displayed at the deepest depth. Aim 3. Integrate ultrasound and optical systems We propose to integrate two ultrasound devices, one therapeutic and the other one diagnostic, on an endoscope that will be used in the esophagus. The therapeutic device will be integrated with the dualaxes confocal microscope, which is introduced through the instrument channel of the endoscope. The therapeutic ultrasound device will be used to enhance delivery of drugs and biomarkers. The relative position of the microscope and the ultrasound therapeutic device will allow real-time monitoring of the ultrasound enhanced delivery of targeted molecular probes and drugs. The diagnostic imaging device will be mounted on the outer surface of the endoscope tube and will provide a 360-degree circumferential image. We propose to build a focused ultrasound transducer on the back side of the parabolic focusing mirror in the dual-axes confocal microscope. The back side of the mirror is a flat surface, circular in shape with a 5-mm diameter and a 2-mm opening in the middle for illumination and collection beams to pass through. A 2-mm diameter ultrasound transducer will be fabricated onto a transparent window substrate providing a transparent ultrasound transducer chip (Fig. 26) that also serves as an optical window. This transparent transducer fits into a 2-mm diameter cut-away section of a catadioptric focusing optic, which provides a parabolic mirror for focusing the dual beams, and a central solid immersion lens for coupling the scanning beam into the tissue. The acoustic focus is designed to coincide with the optical focus of the microscope where the dual beam's axes cross each other. During imaging, this distance varies .between 0 to 0.3 mm away from the back side surface of the combination ultrasound chip/window. In this way, the ultrasound enhanced delivery process can be monitored using the microscope in real time. This transducer chip will be designed to launch a surface acoustic wave (SAW) on the surface that is in contact with the tissue and then take advantage of mode conversion to longitudinal waves to obtain a focused ultrasound beam at the desired location. The waves will propagate in the radial direction inwardly and outwardly. The acoustic energy will leak into the medium in contact at a predetermined angle (by the Snell's law). Due to the circular symmetry of the structure the waves propagating longitudinally in the tissue will focus at the center in the middle of the field-of-view of the microscope. The circular symmetry and the discontinuity at the edges help to bounce the outwardly traveling waves back toward the center to increase the efficiency of the transducer. There are different ways of exciting SAWs on a nonpiezoelectric material. We will build two different types: An interdigital transducer and an edge-bonded transducer.